Investigating the Selective Mechanism of Inhibitors Targeting G Protein-Coupled Receptor Kinase 5 (GRK5) Relative to Its Highly Homologous Counterpart, GRK2

G protein-coupled receptor kinases (GRKs) play a crucial role in mediating various human diseases. However, their high structural similarity poses a significant challenge for developing selective inhibitors. In this study, we conducted molecular docking, molecular dynamics simulations and end-point free energy calculations to understand the binding selectivity of the GRK inhibitor GRL toward GRK5 compared to its highly homologous counterpart, GRK2. Our findings indicate that the driving force behind GRL’s binding selectivity stems from variations in protein–inhibitor van der Waals and electrostatic interactions. Contributions from the phosphate-binding loop and hinge domain are also crucial for determining binding selectivity. Notably, Cys474 within the AST loop of GRK5 forms a covalent bond with the acetamide carbon of GRL, thereby enhancing its specificity. This study provides valuable insights into the development of selective GRK inhibitors.